Elsevier

Behavioural Brain Research

Volume 291, 15 September 2015, Pages 164-171
Behavioural Brain Research

Research report
Endocannabinoid-mediated improvement on a test of aversive memory in a mouse model of fragile X syndrome

https://doi.org/10.1016/j.bbr.2015.05.003Get rights and content

Highlights

  • Deficit on passive avoidance test of memory is a robust phenotype of Fmr1 KO mice.

  • Propofol treatment 30 min after training reverses passive avoidance deficit.

  • Propofol effect blocked by prior treatment with cannabinoid receptor 1 blockade.

  • Treatment with FAAH inhibitor, URB-597, reverses passive avoidance deficit.

  • URB-597 also normalized anxiety-like behavior, but did not enhance social behavior.

Abstract

Silencing the gene FMR1 in fragile X syndrome (FXS) with consequent loss of its protein product, FMRP, results in intellectual disability, hyperactivity, anxiety, seizure disorders, and autism-like behavior. In a mouse model (Fmr1 knockout (KO)) of FXS, a deficit in performance on the passive avoidance test of learning and memory is a robust phenotype. We report that drugs acting on the endocannabinoid (eCB) system can improve performance on this test. We present three lines of evidence: (1) Propofol (reported to inhibit fatty acid amide hydrolase (FAAH) activity) administered 30 min after training on the passive avoidance test improved performance in Fmr1 KO mice but had no effect on wild type (WT). FAAH catalyzes the metabolism of the eCB, anandamide, so its inhibition should result in increased anandamide levels. (2) The effect of propofol was blocked by prior administration of the cannabinoid receptor 1 antagonist AM-251. (3) Treatment with the FAAH inhibitor, URB-597, administered 30 min after training on the passive avoidance test also improved performance in Fmr1 KO mice but had no effect on WT. Our results indicate that the eCB system is involved in FXS and suggest that the eCB system is a promising target for treatment of FXS.

Introduction

Fragile X syndrome (FXS) is the most common inherited intellectual disability and a recognized monogenic cause of autism. FXS is caused by the absence of fragile X mental retardation protein (FMRP) due to silencing of the FMR1 gene. FMRP is an RNA-binding protein that associates with mRNA and stalls ribosome translocation along targeted mRNAs [1]. The absence of FMRP results in a loss of translational control that is thought to be at the core of the disease.

Consistent with this loss of translational control, rates of cerebral protein synthesis (rCPS) measured in vivo are increased in some regions of the brain in an Fmr1 knockout (KO) mouse model of FXS [2]. We attempted to confirm this change in rCPS in the human disease. Surprisingly, our findings indicated that subjects with FXS under propofol sedation had lower rCPS than age-matched, propofol-sedated healthy volunteers [3], despite a lack of an effect of propofol on rCPS in controls [4]. We hypothesized that the decreased rCPS could be due to a selective effect of propofol in subjects with FXS. Further studies in mice showed that propofol had no effect in wild type (WT) mice, but decreased rCPS in Fmr1 KO mice.

In this study, we further investigated this genotype-selective effect of propofol on a test of aversive memory. We studied the effects of propofol on a robust behavioral phenotype in Fmr1 KO mice, deficient performance on the passive avoidance test [5], [6], [7]. It had been reported that in adult, male Sprague-Dawley rats, propofol treatment after training improved performance on this memory test [8]. We assessed the effects of propofol on the passive avoidance test in WT and Fmr1 KO mice, and we report that propofol treatment improved performance in Fmr1 KO, but not in WT mice. Additionally we explored the mechanism by which propofol effects this genotype-specific change in the hope that it might aid in the discovery of new therapeutics. Propofol acts via a wide range of sites, including positive modulation of GABAA receptors [9] and inhibition of fatty acid amide hydrolase (FAAH) [10], an enzyme that catalyzes the metabolism of the endogenous cannabinoid (eCB), anandamide. We focused our study on the latter site of action because it has been implicated in an effect of propofol on the enhancement of memory consolidation in rats [8]. Moreover, eCB systems have been reported to be altered in Fmr1 KO mice [11], [12], [13], [14], [15]. To see if inhibition of FAAH could have therapeutic relevance in FXS, we tested the effects of a specific inhibitor of FAAH on passive avoidance performance and on tests of other behavioral phenotypes in Fmr1 KO mice. Results of our studies point to the eCB system as a promising therapeutic target in FXS.

Section snippets

Animal subjects

Male WT and Fmr1 KO mice on a C57Bl/6J background were bred and genotyped as previously described [2]. Mice were group-housed in a central facility, and naïve mice (90–120 days old) were used for each test. All procedures were performed between 9 am and 11 am and carried out in accordance with the National Institutes of Health Guidelines on the Care and Use of Animals and an animal study protocol approved by the National Institute of Mental Health Animal Care and Use Committee.

Drugs

Propofol

Optimal dose of propofol

We administered propofol (i.p.) 30 min after training on the passive avoidance test at doses of 0, 100, 150, 200, and 300 mg/kg to WT and Fmr1 KO mice (Fig. 1). Following doses of either 100 or 150 mg/kg, mice were lightly sedated. At these doses, mice placed in a supine position could return themselves to a prone position and were fully awake 2.5–3 h after injection. Following a dose of 200 mg/kg, mice were deeply sedated and could not return to a prone position when placed in a supine position. At

Discussion

ECBs act presynaptically to inhibit further release of glutamate or GABA and consequently to modulate activity at mGluR and GABA synapses. Our findings demonstrate that drugs that modulate the eCB system can remedy the deficit in Fmr1 KO mice on a test of long-term memory. We present three lines of evidence: (1) Propofol (reported to inhibit FAAH activity) administered 30 min after training on the passive avoidance test improved performance in Fmr1 KO mice but had no effect on WT. (2) The effect

Acknowledgments

This work was supported by the Intramural Research Program of the National Institute of Mental Health, National Institutes of Health.

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